DESCRIPTION (Abstract of the application) Recombinant adenovirus has been one of the most promising vectors for gene therapy for its ability to infect non-dividing cells and efficient gene delivery in vivo. Recent clinical studies and experiments in animals have shown that the success of adenovirus-mediated gene therapy is limited by two major drawbacks, inflammatory reactions elicited by low levels of viral gene expression in transduced cells and the short duration of therapeutic gene expression. In this application, we propose developing a novel hybrid adeno-EBV vector to overcome the problems associated with the current adenoviral vectors. The hybrid vector is a combination of Generation-X of adenoviral vector which contains no viral genes, and an EBV vector that provides persistent expression of the therapeutic gene. This vector will be replicated in a linear form and packaged into adenoviral particles for efficient gene delivery. Once it enters the target cells, the vector will transform into an EBV vector in a circular form, which can be stably maintained as an episome. A molecular switch, a tet-operon/VP16 system, will be engineered into the vector to regulate the "jumping out" of the EBV vector from the linear adenoviral vector. Once the EBV vector has been excised, the linear portion of the vector which contains the genes coding for the accessory proteins will be degraded as seen in the Generation-X adenoviral vector. This combined vector will have combined advantages of EBV and adenoviral vectors: high titers, efficient transduction (adenovirus), and long-term, high level of gene expression (EBV). In addition, the vector will lack immunogenicity once it has entered the target cells. This will reduce the inflammatory response by the host and prevent the immune-elimination of the transduced cells.